Korean J Physiol Pharmacol.  2012 Oct;16(5):355-360. 10.4196/kjpp.2012.16.5.355.

Simultaneous Determination of Glutamate, Glycine, and Alanine in Human Plasma Using Precolumn Derivatization with 6-Aminoquinolyl-N-hydroxysuccinimidyl Carbamate and High-Performance Liquid Chromatography

• Li QZ
• Huang QX
• Li SC
• Yang MZ
• Rao B

• KMID: 1493970
• DOI: http://doi.org/10.4196/kjpp.2012.16.5.355

Abstract

A simple, sensitive and reproducible high-performance liquid chromatography (HPLC) method has been validated for determining concentrations of glutamate, glycine, and alanine in human plasma. Proteins in plasma were precipitated with perchloric acid, followed by derivatization with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC). Simultaneous analysis of glutamate, glycine, and alanine is achieved using reversed-phase HPLC conditions and ultraviolet detection. Excellent linearity was observed for these three amino acids over their concentration ranges with correlation coefficients (r)>0.999. The intra- and inter-day precision were below 10%. This method utilizes quality control samples and demonstrates excellent plasma recovery and accuracy. The developed method has been successfully applied to measure plasma glutamate, glycine, and alanine in twenty volunteers.

Keyword

Amino acids; AQC; HPLC; Plasma; Ultraviolet detection

MeSH Terms

Alanine
Amino Acids
Aminoquinolines
Carbamates
Chromatography, High Pressure Liquid
Chromatography, Liquid
Glutamic Acid
Glycine
Humans
Perchloric Acid
Plasma
Proteins
Quality Control
Alanine
Amino Acids
Aminoquinolines
Carbamates
Glutamic Acid
Glycine
Perchloric Acid
Proteins

Figure

• 

  Fig. 1 Typical chromatograms of Glu, Gly, and Ala in plasma. (A) Chromatogram for derivatized blank; (B) chromatography for derivatized amino acid standard mixture; (C) representative chromatography of blank (unspiked) pooled human plasma sample; (D) chromatography of the high QC sample (spiked concentrations of Glu, Gly, and Ala were 160, 640, and 800 µM, respectively).

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